Methods of making olive juice extracts containing reduced solids

ABSTRACT

Solids, including fibers can be easily removed from olive juice by mixing the olive juice with a water-miscible solvent to form two phases and separating the phases. Preferably the solvent is ethanol.

CROSS-REFERENCE

This application is a divisional of commonly owned co-pending U.S.Application Ser. No. 12/674,044 filed Feb. 18, 2010, which is thenational phase under 35 USC §371 of International Application No.PCT/EP2008/006790 filed Aug. 19, 2008, which designated the U.S. andclaims priority to Europe Application No. 07016344.9 filed Aug. 21,2007, the entire contents of each of which are hereby incorporated byreference.

FIELD OF THE INVENTION

This invention relates to olive juice extracts containing an decreasedamount of solids, and to methods of making these extracts. Olive juiceextracts can be used as nutritional supplements, as they have potentantioxidant activities. Nutritional compositions containing these novelextracts are also part of this invention.

BACKGROUND OF THE INVENTION

The medicinal benefits of olive oil and olive extracts is widely beingrecognized. To make olive oil, the olive fruits are ground into a paste.Pressure is applied to the paste to separate the oil from the groundfruit. In addition to providing olive oil, the pressing also releasesthe water content of the olive fruit, which contains many water solublephytochemicals. This water is known by a number of names, including“vegetation water, olive juice, and olive waste water”. Interestingly,while olive juice and its disposal can be a problem for the olive oilproducer, this olive juice can be a desirable rich source of phenoliccompounds, which can have beneficial nutritional properties.

In the past, methods to concentrate the nutritional olive juice involvedtime consuming incubation, filtration and/or centrifugation, and/orspray drying steps. Another problem is that the usability of dried orliquid olive juice in food or dietary supplements is limited due to thesmell, bitterness, and turbidity of the olive juice, as well as the lowcontent of hydroxytyrosol, one of the active polyphenols.

It would therefore be desirable to develop a better method for producingan all-natural, hydroxytyrosol-rich, non-bitter olive juice extractwhich is efficient and cost-effective.

DESCRIPTION OF THE INVENTION

In accordance with this invention, new processes for making anall-natural, hydroxytyrosol-rich, non bitter olive juice extract arepresented. Also as part of this invention are novel juice extract, andcompositions containing this novel olive juice extracts. Thus thisinvention relates to a method of removing at least some of the solidspresent in an olive juice comprising: a) adding a water-miscible ornearly water-miscible solvent in an amount (by volume) equal to about40-400% of the olive juice to form two phases; and b) separating thephases.

A typical olive will contain approximately 50% water, 22% oil, 19%carbohydrates, 6% cellulose, 2% proteins, and oleuropein andhydroxytyrosol (combined) 0.2%. It should be appreciated that the exactmakeup of the fruit (and its subsequent extract) will vary according tothe variety of olive used, the time of harvest, and even growingconditions.

Contrary to literature reports (see e.g., Briante et al, 2002, J.Biotechn. 93: 109-119, and Soler-Rivas et al 2000, J Sci Food Agric80:1013-1023) it has been found, in accordance with this invention, thatthe unknown compounds which impart a bitter taste to the olive juice areneither hydroxytyrosol nor oleuropein. While not wishing to be bound bytheory, they may contain a labile phenolic ester group. Regardless ofthe identity of the bitter compounds, they are very sensitive to base,and are not stable at higher pH.

As used throughout the specification and claims, the followingdefinitions apply:

“HT” means hydroxytyrosol.

“Olive Juice”, “Olive Waste Water” and “Vegetation Water” are terms allintended to be used interchangeably. They refer to the water phaseproduced during olive oil production. It is a slurry with a complexmixture of carbohydrates along with the compounds of interest, such asHT and oleuropein (which contains bound HT, and which may besubsequently broken down to yield HT).

A. Simultaneously Removing The Bitter Taste and Increasing The Amount ofHT

The following step is an optional, but preferred step in the practice ofthis invention. Reference is made to co-pending U.S. application Ser.No. 12/674,213 filed on Jun. 6, 2011. If desired, the process can beginwith the procedures described in Section B, below.

Any form of olive juice may be used as a starting material for theprocesses of this invention, although minor variations of the processesmay be required for optimal results. Fresh juice typically containingabout 85-90% water and 10-15% solid matter can be purchased and used asa starting material in the processes of this invention. Other forms ofcommercially available olive juice which have been filtered by thesupplier to remove some or all of the solids, are also appropriatestarting materials, as well. Also appropriate is olive juice which ispurchased in a concentrated form (for example, 3-4×), or concentratedprior to use by simple evaporating techniques known in the art.

In other cases, the olive juice may be conveniently purchased in astabilized, freeze dried form where citric acid has been added tostabilize it, for example, HIDROX from CreAgri (California, USA). Driedolive juice typically contains about 60% carbohydrates, 10% fibres, 10%fat, and 6% polyphenols (this includes approximately 2% hydroxytyrosol,0.2% tyrosol and other polyphenols), and organic acids, proteins andminerals. To use this dried form as a starting material for thisinvention, it should be hydrated. The resulting juice is a preferredstarting material.

It has been found, in accordance with one aspect of this invention, thatthe polyphenol concentration of any type of olive juice can be increasedby maintaining the olive juice at an elevated temperature and increasingthe pH to at least 6.

The elevated temperature can exist over a wide range, i.e. from about 20degrees C. up to about 100 degrees C. The actual temperature is notespecially critical, as long as it is under the boiling point of theolive juice. Preferred temperature ranges are from about 60 to about 80degrees C., and a more preferred temperature is about 80 degrees C.

Along with the increase in the temperature, the pH of the juice shouldbe adjusted to a higher pH than is originally present in the untreatedolive juice. The adjusted pH should be in a range of about 6 to about11, with the preferred pH being from about 7 to about 10, and a morepreferred range is from about 8 to about 9. A pH above about 10 canresult in the degradation of the final HT, so it is not preferred. Anybasic compound capable of adjusting pH can be used: preferred compoundsare NaOH, KOH, and mixtures thereof. Generally, the amount needed willvary from batch to batch, but a typical amount needed will beapproximately 10-15% by weight based on the weight of the batch.

Under the conditions of the elevated temperature and pH-complexcompounds which contain bound HT may break down, liberating HT. Thus,after the reactions are substantially complete, the juice will contain ahigher amount of HT than the starting material. During the time that thereactions are taking place, the pH may change, and re-adjustment(s) maybe required to maintain the desired pH. It is preferred to continuallyadjust the pH during the time of the reaction. One way of determiningthat the reactions are substantially complete is that the pH will remainrelatively stable (i.e. relatively unchanged) for at least about tenminutes. The amount of time it will take for a batch to reach the pointof stability for about 10 minutes can vary greatly depending on theindividual composition to of the batch. Generally, as higher thetemperature and the pH the less time the reaction will take.

At this point, the partially processed olive juice no longer is bitter.Thus this invention also includes a process for removing the bitternessfrom an olive juice extract comprising raising the temperature and pH ofthe olive juice to the above-mentioned conditions. At this point, theolive juice may be process as desired, using any of the known processsteps in the art, or, optionally steps presented below.

B. Removal of Solids and Fibers

The starting material for this step may be the de-bittered, HT enhancedjuice made in accordance with the above description A, or it may be anystarting material (such as those described in Section A as suitable asstarting materials for the optional step described there. Generallyconcentrated juice is preferred for economic reasons, but the methodwill work with a variety of juices. Thus, the starting material may havesolids, fats (oils) and/or fibers present in it.

Typically unprocessed olive juice will contain a large amount of finefibers. These are difficult to filter out as they tend to clog membranefilters, requiring a large amount of maintenance. It has been found, asanother aspect of this invention, that fibers, residual oil and lipids,and/or other solids present in the olive juice can be agglomerated andthen easily removed by a) adding a solvent in an amount (by volume)equal to about 40-400% of the olive juice or partially concentratedolive juice to form two phases; and then b) separating the two phases.

The solvent which can be employed in this process can be any solvent ormixture of solvents which is water-miscible or nearly miscible liken-butanol. Alcohols, especially C1 to C4 (or mixtures thereof) arepreferred, and ethanol is particularly preferred, especially when theend product is to be used in food or as part of a medicinal ornutritional supplement product. Other solvents which can also beutilized include: acetonitrile, acetone, and glycol.

The amount of solvent is not especially critical, and will depend on theamount of water in the mixture. Usually the solvent will be needed in anamount that is at least equal to the amount of water present. Forexample, if ethanol is the solvent, for every 500 ml of water, about 600ml of EtOH will be required. However, this is subject to a wide range:from about 40-400% solvent compared to the amount of water by weight,preferably 100-240%, and more preferably about 160-220% solvent comparedto the amount of water. The important criteria is that enough solvent isadded so that two distinct phases are formed.

The temperature of this step is not particularly critical and may rangefrom 0 degrees Celsius to about 80 degrees Celsius, or any temperaturewhich is less than the boiling point of the mixture. Conveniently thetemperature will be between about 20 and 60 degrees C., and mostconveniently is room temperature, or about 25 degrees.

At this point, two phases exist and any convenient means can be used toseparate the two phases. The method of separation will depend on theactual make up of the material. For some batches, a filter can now beeasily used, and/or a centrifuge. Other batches could be easilyprocessed by simple decantation (if not too many solids were present, orif material is stuck to the wall of the vessel) or ordinary phaseseparation.

In another embodiment of this invention, this same process can be usedwith a starting olive juice which has been processed to remove fibers,but which has undergone the de-bittering/HT concentrating step describedin the previous section. Addition of the solvent and separation of theresultant phases will allow separation of any precipitates which mayhave formed during the debittering process.

C. Recovery of the Solvent

The solvent can be recovered for re-use, if desired, making the processmore economical. Evaporation is generally the easiest way to so recoverthe solvent. At this point, is has been found in yet another aspect ofthe invention, that if one chooses to employ an evaporation step torecover the solvent, then adjusting the pH of the extract to below 6,preferably below 5, will prevent unwanted foaming during the evaporationprocess.

At this point, the resulting non-bitter, solids-free olive juiceextract, which forms another aspect of this invention can be used as isin any way desired. For example, it can be directly incorporated into anutritional composition, for example a food or beverage composition. Thefood can be suitable for humans consumption, or can be an animal feed.Alternatively, it can be made into a nutritional supplement, forexample, it can be formulated into capsules, tablets, or the like usingknown methods. Alternatively, the composition can be used in cosmeticcompositions.

In other embodiments of this invention, the non-bitter, solids-freeextract is spray dried or free-dried using conventional techniques toform a powder derivative, and then incorporated into a final product.Optionally, stabilizers and the like amy be added to the juice prior tothe drying step. The resulting powder can be directly incorporated intoa nutritional composition, or it can be an animal feed. Alternatively,it can be made into a nutritional supplement; for examples it can beformulated into capsules, tablets, or the like using known methods.Alternatively, the composition can be used in cosmetic compositions.

In other embodiments of this invention, the non-bitter, solids-freeextract is subjected to additional processing steps, such as de-fatting,further extraction, or distillation of the HT. Such processes may beknown techniques, or the techniques as further described in co-pendingpatent application U.S. application Ser. No. 12/647,032 filed on Feb.18, 2010 may be used.

The invention is now further illustrated in the following non-limitingexamples.

EXAMPLES Example 1 Starting Material

Unless otherwise stated, all % s given refer to weight percent.

Starting material was HIDROX 6% (from CreAgri, Haywood, Calif. Itscontent is: (in Lot 6022406002. This is freeze-dried olive juice whichhas been stabilized with citric acid. weight %):

Carbohydrates Approx. 60% Lipids (olive oil) Approx. 15% Fibers Approx.10% Hydroxytyrosol 2.1% Water Approx. 2%

Appearance:

Solubility in water: a turbid, fine suspension Taste bitter Colorbeige-brown powder

Example 2 Agglomeration of Solids with Ethanol

100 g HIDROX (CreAgri, from Example 1) 6% polyphenols, and 80 ml waterwere added to a reaction flask in a nitrogen atmosphere. The mixture waswarmed up to 80° C. and adjusted to pH 9.0, followed by stirring for 30minutes at 80° C. at pH 9.0 with continuous addition of a total of 38 mlsodium hydroxide 10 mol/l to maintain pH. The suspension was cooled to60° C.

The content of hydroxytyrosol increased by the base treatment to approx.130%.

The solids where precipitated at 60° C. by addition of 200 g ethanol.The mixture was cooled, stirred at ambient for 30 minutes and filteredby a Buchner funnel. The filtercake was washed with 50 g of ethanol70%/water 30%.

The filtrate was adjusted to pH 5 with approx. 5 ml hydrochloric acid 10mol/l and evaporated at the rotavapor (20 mbar, 60° C.) to recover theethanol, the non volatile residue 33.5 g.

Content [w %]: 7.4% hydroxytyrosol, 0.8% tyrosol.

Yield: 119% of the initial hydroxytyrosol

The extract is not bitter at 1 mg/ml water

Example 3

In Table 1, below, “yield %” refers to the content of HT in the driedfiltrate [w %]. Yield [w %] is based on the initial content of thestarting material, HIDROX 6%.

TABLE 1 Ethanol Needed for the agglomerations of solids from hydrolyzedolive juice Agglomeration HT in filtrate pH with ethanol [m]* Filtrationw % yield % 3.5 ~3 slow 4.2% 132% 5.0 ~2.3 rapid 7.2% 128% 6.5 ~1.1rapid** 8.9% 126% 8.0 ~0.6 rapid** 8.1% 108% *Agglomeration was atambient of a 50% solution of HIDROX (see Ex. 1) in water hydrolyzed at80° C. pH = 9. pH was adjusted with conc. HCl at RT. [m] = amounts ofethanol: 50% solution.[w/w] Filterability and coagulation can beadjusted by the amount of ethanol (optimum different for each pH. Thedried precipated mass contained 0.1-0.4% HT

Example 4

100 g HIDROX 6% polyphenols (CreAgri, Example 1) and 80 ml water wereadded to a reaction flask in an inert gas atmosphere. The mixture waswarmed up to 60° C. under a nitrogen blanket and adjusted to pH6.0followed by stirring for 30 minutes at 60° C. at pH 6.0 with continuousaddition of a total of 36 ml sodium hydroxide 10 mol/l to maintain pH.

The content of hydroxytyrosol increased by the base treatment to approx.115%.

The solids where precipitated at 60° C. by addition of 240 g ethanol.The mixture was cooled, stirred at ambient for 30 minutes and filteredby a Buchner funnel. The filtercake was washed with 50 ethanol 70%/water30%

The filtrate was evaporated at the rotavapor (20 mbar, 60° C.) torecover the ethanol. We recovered: 33.5 g residue. Content [w %]: 6.3%hydroxytyrosol, 0.7% tyrosol. The residue is slightly bitter at 1 mg/mlwater

Example 5

The following samples where made in accordance with the above Examples

100 g HIDROX 6% (starting material, Example 1) was mixed with 100 gwater to give a 50% solution. pH was adjusted to pH 9 at 80° C. with 10m sodium hydroxyde. After approximately 30 minutes at 80° C.,hydroxytyrosol which was in bound form is liberated, and the yieldincreases (theoretical yield is approx. 135%). With increasing pH, lessethanol is needed to agglomerate the particles, as described in theTable, below. Too little ethanol will not agglomerate the particlesenough to precipitate efficiently to form a easily removable part.Higher amounts of ethanol results in a higher yield. The highestconcentration of hydroxytyrosol in the filtrates resulted at a pH 8-9.For a 50% solution of olive juice, about the same amount of ethanolseems optimal.

Hydrolysis pH pH 3.5 pH 8.0 pH 8.0 pH 9.0 pH 9.0 temp 60° C. 80° C. 80°C. 80° C. 80° C. Agglomeration pH pH 8 pH 6.5 pH 8 pH 8 pH 8 Ethanol 120 g 188 g 210 g  165 g 215 g RT 60° C. 60° C. RT 60° C. Soluble 36.4g 32.0 g 32.9 g 33.9 g 31.6 g Residue [g] HT [w %] 5.34% 7.17% 7.24%7.22% 7.64% Yield [w %] 92.6%  109%  113%  117%  115% Bitterness atBitter Sl. Not Not Not 1 mg/ml bitter bitter bitter bitter

Example 6

243 g of a commercially available olive juice extract which has beenprocessed by the to provider to remove fibers (content=0.89%hydroxytyrosol) was concentrated in a vacuum to 190 g. In a reactionflask under an atmosphere of nitrogen, the concentrate was heated to 80°C. and pH was adjusted to 8.0, followed by stirring for 30 minutes at80° C. at pH 8.0 with continuous addition of a total of 24 ml sodiumhydroxide 10 mol/l to maintain pH.

To the dark colored mixture was added 280 g ethanol to make theprecipitate. The mixture was cooled and stirred at ambient for 1 h. Theupper solution was decanted from the precipitate and evaporated at therotavapor to recover the ethanol. The nonvolatile residue was 78 g.Content [w %]: 2.6% hydroxytyrosol, 0.6% tyrosol. Yield: 93% of theinitial hydroxytyrosol. The residue is not bitter at 1 mg/ml water.

1. A non-bitter hydroxytyrosol enriched olive juice extract produced byremoving at least some of the solids present in an olive juice by amethod consisting essentially of: (a) adding a pH adjuster to the olivejuice to obtain a pH of at least 6; (b) adding to the olive juice atleast one water-miscible or nearly water-miscible solvent selected fromthe group consisting of methanol, ethanol, propanol, butanol,acetonitrile, acetone and glycol in an amount equal to about 40-200% byvolume of the olive juice to form two phases; and (c) separating the twophases and recovering the phase containing an olive juice which has atleast some of the solids removed.
 2. The olive juice extract accordingto claim 1, wherein the solvent is ethanol.
 3. The olive juice extractaccording to claim 1, wherein step (c) comprises at least one stepselected from the group consisting of a filtering step, a centrifugationstep, a decanting step, and a step of removing a layer of the twophases.
 4. The olive juice extract according to claim 1, wherein themethod further consisting essentially of (d) recovering the solvent ofstep (b).
 5. The olive juice extract according to claim 4, wherein thestep (d) is practiced by evaporating the solvent while maintaining a pHbelow
 6. 6. The olive juice extract of claim 1 or 4, wherein step (a) ispracticed by maintaining the olive juice at an elevated temperature andincreasing the pH to at least 6 by adding the pH adjuster until thepolyphenol concentration of the olive juice is increased.
 7. The olivejuice extract according to claim 1, wherein the solids present in olivejuice are not fibres.
 8. The olive juice extract according to claim 6,wherein the elevated temperature is from about 20 degrees to 100 degreesC., and the pH is from about 6 to about
 10. 9. The olive juice extractaccording to claim 8, wherein the elevated temperature is maintaineduntil the pH remains unchanged for at least about 10 minutes.
 10. Theolive juice extract according to claim 6, wherein step (a) is practicedby adding NaOH, KOH or mixtures thereof as the pH adjuster to the olivejuice.
 11. The olive juice extract according to claim 9, wherein thetemperature is about 60-90° C. and the pH is about 7-9.
 12. The olivejuice extract of claim 1, wherein the method further consistingessentially of (d) freeze drying or spray drying the olive juice. 13.The olive juice extract of claim 4 or 12, wherein step (c) comprises atleast one step selected from the group consisting of a filtering step, acentrifugation step, a decanting step, and a step of removing a layer ofthe two phases.
 14. The olive juice extract of claim 12, wherein step(a) is practiced by maintaining the olive juice at an elevatedtemperature and increasing the pH to at least 6 by adding the pHadjuster until the polyphenol concentration of the olive juice isincreased.
 15. The olive juice extract of claim 12, wherein the solidspresent in the olive juice are not fibres.
 16. The olive juice extractof claim 15, wherein the elevated temperature is from about 20 degreesto 100 degrees C., and the pH is from about 6 to about
 10. 17. The olivejuice extract of claim 16, wherein the elevated temperature ismaintained until the pH remains unchanged for at least about 10 minutes.18. The olive juice extract of claim 14, wherein step (a) is practicedby adding NaOH, KOH or mixtures thereof as the pH adjuster to the olivejuice.
 19. The olive juice extract of claim 17, wherein the temperatureis about 60-90° C. and the pH is about 7-9.
 20. The olive juice extractof claim 1, comprising at least about 2-8% hydroxytyrosol, other olivepolyphenols, and no solids.
 21. The olive juice extract of claim 20which is in powdered form.
 22. A nutritional composition comprising theolive juice extract of claim
 1. 23. A feed composition comprising theolive juice extract of claim
 1. 24. A cosmetic composition comprisingthe olive juice extract of claim 1.